The long-term objective of this research is the development of rational methods for the design and improvement of membrane-penetrating, amphipathic peptides that are antibiotic or cytolytic, or can carry other drug molecules as cargo into cells. Critical knowledge in reaching this objective is the determination of the mechanism of membrane penetration by these peptides. Peptides in this class are known to exhibit considerable target specificity, which appears to derive from the interaction of the peptides with the lipid bilayer of the target cell membrane without the intervention of protein receptors. Furthermore, the development of resistance by bacteria is much more difficult because it entails massive changes in the bacterial membranes. Three fundamental hypotheses relating to the mechanism of these peptides will be tested. The prediction is that specific features of the peptide sequences are necessary for peptides to penetrate cells or disrupt the membrane. Four naturally-occurring peptides were selected as the basis for new sequences that will be used to test the hypotheses. If correct, a powerful predictive program will be available to design peptides that function with a desired mechanism. Thus, the aim is to design peptides for cargo-delivery into cells or for cytolytic functions. Cargo-carrying peptides can be used to transport drugs into eukaryotic cells or antibiotics into bacterial cells. Surmounting cellular barriers, including intracellular compartments, is a major difficulty in the use of antibiotics. Furthermore, as bacteria increasingly develop resistance against conventional antibiotics, understanding the physical properties necessary for the rational design of new antibiotics that are not prone to resistance development is of the utmost importance.